Project Summary Age-related macular degeneration is a principal cause of vision loss in individuals over the age of 60 years including Veterans. It is characterized by a loss of sight in the central visual field where sharp, polychromatic images are generated under the bright light conditions that modern humans are typically exposed to during waking hours. Loss of this high acuity color vision leads to significant disability. The high prevalence of AMD places a large burden on the healthcare system with upwards of 98 billion dollars spent yearly on AMD-related healthcare costs in the US. Currently, there are no highly effective treatments for the most common form of the disease, known as geographic atrophy, which makes up ~90% of advanced AMD. During the past VA funding period, we studied inhibition of the visual cycle as a potential treatment for AMD and characterized enzymes proposed to contribute to the normal function of the visual cycle pathway. Based on key findings we made during the initial funding period, we now propose to investigate related pathways that are tied to the pathogenesis of AMD through the involvement of retinaldehyde (RAL) derived from the visual cycle. Additionally, we will continue our studies on an enzyme known as Des1 that has been proposed to mediate the regeneration of cone visual pigments, the color sensing molecules in the central retina. We will explore these pathways through the following Specific Aims: 1. Elucidate biological roles for Des1 within the RPE using novel RPE- specific Cre mice. Previously, we showed that Des1 protein in Mller cells is not a major contributor to 11-cis- retinol synthesis for cone photoreceptors. However, single cell RNA-Seq analysis revealed that the RPE is the principal site of Des1 expression in the retina raising questions about its biological role in this tissue. Using a validated floxed Des1 mouse model, we will investigate the impact of Des1 loss of function on RPE and photoreceptor health and visual cycle function by crossing these mice with RPE-specific Cre mice and characterizing them using a variety of functional and imaging techniques. Des1 also plays a key role in de novo ceramide production, a known mediator of apoptotic RPE cell death. We hypothesize that Des1 deletion in the RPE will modulate susceptibility of the tissue to chemical-induced toxicity, which serves as a model for RPE cell death that occurs in geographic atrophy. These studies will test the viability of Des1 as a potential target for AMD therapeutics. 2. Advance next-generation visual cycle modulators (VCMs) with selective pharmacodynamics. Visual cycle modulators were originally designed to inhibit RPE65 in order to suppress pathological lipofuscin accumulation and slow retinal disease progression. We discovered a novel mechanism of action for these compounds: direct reaction with RAL released from activated visual opsins to limit formation of pathological RAL adducts. We have generated visual cycle modulators with preferential activity towards RAL sequestration that possess protective effects against retinopathy with reduced effects on visual cycle activity. Based on our initial studies, we propose to synthesize and characterize a new set of rationally-designed visual cycle modulators that we hypothesize will possess augmented therapeutic activity and diminished visual cycle suppression. 3. Develop phosphodiesterase (PDE) inhibition as a treatment for retinal disease. Prior research has implicated aberrant GPCR signaling in RAL toxicity. Phosphodiesterase enzymes are major effectors and regulators of GPCRs and have been successfully targeted for clinical applications. We hypothesize that inhibitors of PDEs will confer protective effects against retinal insults without impairing visual function. Our preliminary data indicates that PDE4 inhibitors are particularly effective at low doses in animal models of RAL toxicity. We will screen these compounds and related derivatives to elucidate their site and mechanism of protective action using animal models of retinopathy. Together, these studies may uncover small molecules that could readily be translated into retinal disease treatments for veterans.